Activation of cell-surface receptors releases inside cells Ins(1,4,5,)P3 which mobilizes cellular Ca2+ stores as part of a ubiquitous signalling cascade. This project studies the enzymes metabolizing Ins(1,4,5,)P3, since this deactivates signalling, while introducing down-stream metabolites such as Ins(1,34,5)P4 that may themselves have key functions. We have purified and characterized an active Ins(1,3,4,5)P4 3-phosphatase and shown both this activity, and Ins(1,4,5)P3 5- phosphatase, to be perturbed in some cases of cell transformation, and following HIV infection of lymphocytes. We are pursuing the mechanisms of these effects. Our demonstration that the 3-phosphatase is compartmentalized in the lumen of endoplasmic reticulum has led us to pursue its possible involvement in protein trafficking. We are currently attempting to sequence this enzyme. With the aim of understanding the physiological significance and mechanisms behind receptor-regulated InsP5 turnover (itself a novel effect recently uncovered in this laboratory), three major enzymes of InsP5 synthesis have also been highly purified: Ins(3,4,5,6)P4 1-kinase, Ins(1,3,4,)P3 6-kinase, and Ins(1,4,5,6)P4 3-kinase. With an underlying goal of studying the impact of environmental toxins on signalling processes, we have made progress in elucidating the mechanisms by which aluminum toxicity may manifest itself; although our work shows no direct effect upon Ca2+ signalling, we have discovered this ion may redirect the synthesis of the cell's complement of InsP5 and InsP6. We have also discovered that low concentrations of fluoride can regulate the interconversion of inositol polyphosphates with a novel metabolic group of inositol pyrophosphates. We have also obtained evidence that levels of these inositol pyrophosphates are profoundly affected by the toxin thapsigargin.